Recently, significant progress has been made in the methodology for the rapid determination and analysis of DNA sequence. However, significant gains in methodology need yet be made before sequencing of the human genome will be aggressively pursued. By contrast, we propose that current methodology is significantly advanced to be of great use in the analysis of prokaryotic genomes, which are commonly 1-5 megabases (Mb) in size. Determining the nucleotide sequence of a prokaryotic genome will dramatically enhance the rate at which we will come to understand the genetic basis for the biology in which that organism participates. This endeavor will also be extremely cost-effective. The NIH has a considerable interest in understanding the pathogenesis of many human bacterial infections. Therefore, we propose to examine a human bacterial pathogen to test whether genomic sequencing will enhance microbial pathogenesis studies. We have selected Neisseria gonorrhoeae as this model organism, for several reasons. Most importantly recent data indicates that N. gonorrhoeae population structure is panmictic. in contrast to several other pathogens whose populations have a clonal structure. Selecting an organism from a panmictic population ensures that we will not pick the wrong strain for sequencing, as all strains appear of equivalent virulence. N. gonorrhoeae (the gonococcus) is the causative agent of human gonorrhea, a sexually transmitted disease with significant impact on the health and well-being of the U.S. populace. Typically, 1 million or more gonococcal infections are reported in the U.S.; this is considered an underestimate, due to underreporting. Gonococcal disease costs the U.S. health care system over $3 billion annually. Although gonococcal infections are often trivialized by the lay public, the organism can cause severe disease, particularly in women. Infections in women are often asymptomatic and undiagnosed until complications (pelvic inflammatory disease and salpingitis) appear, causing significant and often permanent damage to the female reproductive tract. Antibiotic resistance is also of increasing concern. Consequently, the NIH has devoted significant resources for understanding and controlling gonococcal infections. We propose to determine the nucleotide sequence of N. gonorrhoeae strain FA109O; for several reasons, FA109O is the optimal strain for these studies. This study is part of IRPG that includes a companion application to determine the genome sequence of Streptococcus pyogenes. The assembly of scientists and varied expertise in microbial pathogenesis and genomic sequencing will produce a synergy that will provide new insights and reagents essential for the study of these medically important microorganisms.